Engagement of the BCR increases glucose uptake in a PI-3K–dependent pathway. (A) Splenic B cells were cultured in the absence (Med) or presence of 10 μg/mL anti-Ig (Ig) for 15 hours. Uptake of 2-[³H]deoxyglucose was measured for 60 seconds in the presence or absence of 10 μM cytochalasin B. Transport represents the difference between the total 2-[³H]deoxyglucose uptake (cpm/10⁶ cells/sec) minus uptake in the presence of 10 μM cytochalasin B. (Inset) Splenic B cells were cultured in the presence of 10 μg/mL anti-Ig (Ig) for 15 hours. Cells were collected, and the initial uptake of 2-[³H]deoxyglucose was measured for the indicated times (in seconds) in the presence (bottom line) or absence (top line) of 10 μM cytochalasin B as described in “Materials and methods.” The data indicate that uptake of 2-[³H]deoxyglucose in anti-Ig–stimulated B cells is linear for 90 seconds. (B) Parallel B cells were also stimulated with 10 μg/mL anti-Ig (15 hours) in the absence (Ig) or presence of 10 μM LY294002 (Ig+LY), 50 nM wortmannin (Ig+Wort), or 20 nM rapamycin (Ig+Rap). Cells were then harvested, and 2-[³H]deoxyglucose uptake was measured. The inhibitors had no measurable effect on 2-[³H]deoxyglucose uptake in B cells cultured in medium alone (data not shown). Error bars reflect standard deviation from the mean of triplicate measurements, and the data for panels A-B are representative of 3 independent experiments. (C) B cells were pretreated with 10 μM LY294002 (LY), 50 nM wortmannin (Wort), or 20 nM rapamycin (Rap) for 30 minutes and cultured in medium alone (M) or stimulated with 10 μg/mL anti-Ig (αIg) for 15 minutes. Cell lysates were prepared, and phosphorylation of Akt on Ser473 and p70S6K on Thr389 was monitored by Western blot.²¹  The data are representative of 4 independent experiments.